Isolation of T‐Tubules from Skeletal Muscle

Zorzano, António; Camps, Marta

doi:10.1002/0471143030.cb0324s31

Cited by 7 publications

(12 citation statements)

References 34 publications

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“…Isolation of T-tubule-enriched membrane fractions was done essentially as described (38). Similar results were obtained in replicate experiments using mice both with and without a GLUT4myc transgene (kindly provided by Dr. Amira Klip; Ref.…”

Section: Methodsmentioning

confidence: 73%

Enhanced Fasting Glucose Turnover in Mice with Disrupted Action of TUG Protein in Skeletal Muscle

Löffler¹,

Birkenfeld²,

Philbrick³

et al. 2013

Journal of Biological Chemistry

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Background: Insulin stimulates endoproteolytic cleavage of TUG proteins to promote glucose uptake in cultured adipocytes, but the role of this pathway in muscle is uncharacterized. Results: Transgenic mice with constitutive and unregulated TUG cleavage in muscle had increased whole-body and musclespecific glucose uptake during fasting. Conclusion: Insulin acts through TUG to control glucose uptake in muscle. Significance: Understanding insulin action will elucidate diabetes pathogenesis.

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Section: Methodsmentioning

confidence: 73%

Enhanced Fasting Glucose Turnover in Mice with Disrupted Action of TUG Protein in Skeletal Muscle

Löffler¹,

Birkenfeld²,

Philbrick³

et al. 2013

Journal of Biological Chemistry

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“…Another would be pre-stretching of the cardiac tissue prior to cell isolation, as this can cause surface membrane incorporation of caveolae 43 . Thirdly, one could isolate the T-tubules using sequential centrifugation of homogenised cardiomyocytes followed by purification of T-tubule membranes by vesicle immuno-isolation and reconstitution into a continuous membrane 136 . It might then be possible to directly patch clamp SAC on the isolated T-tubule membrane.…”

Section: Discussionmentioning

confidence: 99%

Molecular candidates for cardiac stretch-activated ion channels

Reed

Kohl

Peyronnet

2014

Global Cardiology Science and Practice

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The heart is a mechanically-active organ that dynamically senses its own mechanical environment. This environment is constantly changing, on a beat-by-beat basis, with additional modulation by respiratory activity and changes in posture or physical activity, and further overlaid with more slowly occurring physiological (e.g. pregnancy, endurance training) or pathological challenges (e.g. pressure or volume overload). Far from being a simple pump, the heart detects changes in mechanical demand and adjusts its performance accordingly, both via heart rate and stroke volume alteration. Many of the underlying regulatory processes are encoded intracardially, and are thus maintained even in heart transplant recipients. Over the last three decades, molecular substrates of cardiac mechanosensitivity have gained increasing recognition in the scientific and clinical communities. Nonetheless, the processes underlying this phenomenon are still poorly understood. Stretch-activated ion channels (SAC) have been identified as one contributor to mechanosensitive autoregulation of the heartbeat. They also appear to play important roles in the development of cardiac pathologies – most notably stretch-induced arrhythmias. As recently discovered, some established cardiac drugs act, in part at least, via mechanotransduction pathways suggesting SAC as potential therapeutic targets. Clearly, identification of the molecular substrate of cardiac SAC is of clinical importance and a number of candidate proteins have been identified. At the same time, experimental studies have revealed variable–and at times contrasting–results regarding their function. Further complication arises from the fact that many ion channels that are not classically defined as SAC, including voltage and ligand-gated ion channels, can respond to mechanical stimulation. Here, we summarise what is known about the molecular substrate of the main candidates for cardiac SAC, before identifying potential further developments in this area of translational research.

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“…This protocol is based on previously reported methods used in triad isolation from rabbit muscle membranes with slight modifications (Saito et al, 1984;Zorzano and Camps, 2006). Twelve grams of rabbit skeletal muscle were minced into 2-4 mm 2 pieces and fat and tendon were removed.…”

Section: Muscle Fractionationmentioning

confidence: 99%

“…Muscle fractions were separated by SDS/PAGE gel, and validated using several protein markers for SR, T-tubule and sarcolemma. The caveolin 1 and actin was used as loading control as they are associated to both fractions (Zorzano and Camps, 2006). Further, we assessed the presence of MTM1 in F3+F4 fractions using the vesicle immunoisolation technique (Horgan and Kuypers, 1988;Rosemblatt et al, 1981).…”

Section: Muscle Fractionationmentioning

confidence: 99%

Myotubularin and PtdIns3P remodel the sarcoplasmic reticulum in muscle in vivo

Amoasii

Hnia

Chicanne

et al. 2013

Journal of Cell Science

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SummaryThe sarcoplasmic reticulum (SR) is a specialized form of endoplasmic reticulum (ER) in skeletal muscle and is essential for calcium homeostasis. The mechanisms involved in SR remodeling and maintenance of SR subdomains are elusive. In this study, we identified myotubularin (MTM1), a phosphoinositide phosphatase mutated in X-linked centronuclear myopathy (XLCNM, or myotubular myopathy), as a key regulator of phosphatidylinositol 3-monophosphate (PtdIns3P) levels at the SR. MTM1 is predominantly located at the SR cisternae of the muscle triads, and Mtm1-deficient mouse muscles and myoblasts from XLCNM patients exhibit abnormal SR/ER networks. In vivo modulation of MTM1 enzymatic activity in skeletal muscle using ectopic expression of wild-type or a deadphosphatase MTM1 protein leads to differential SR remodeling. Active MTM1 is associated with flat membrane stacks, whereas deadphosphatase MTM1 mutant promotes highly curved cubic membranes originating from the SR and enriched in PtdIns3P. Overexpression of a tandem FYVE domain with high affinity for PtdIns3P alters the shape of the SR cisternae at the triad. Our findings, supported by the parallel analysis of the Mtm1-null mouse and an in vivo study, reveal a direct function of MTM1 enzymatic activity in SR remodeling and a key role for PtdIns3P in promoting SR membrane curvature in skeletal muscle. We propose that alteration in SR remodeling is a primary cause of X-linked centronuclear myopathy. The tight regulation of PtdIns3P on specific membrane subdomains may be a general mechanism to control membrane curvature.

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Isolation of T‐Tubules from Skeletal Muscle

Cited by 7 publications

References 34 publications

Enhanced Fasting Glucose Turnover in Mice with Disrupted Action of TUG Protein in Skeletal Muscle

Enhanced Fasting Glucose Turnover in Mice with Disrupted Action of TUG Protein in Skeletal Muscle

Molecular candidates for cardiac stretch-activated ion channels

Myotubularin and PtdIns3P remodel the sarcoplasmic reticulum in muscle in vivo

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